# US 10,246,500 B2 · Human cytolytic fusion proteins > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 10,246,500 B2 - Document ID: US10246500B2 - Status: Granted - Kind: B2 - Application number: 15/318,584 - Filed: 2015-06-15 - Published: 2017-04-27 - Granted: 2019-04-02 - Earliest claimed: 2014-06-13 - Assignees: University of Cape Town - Applicants: University of Cape Town - Inventors: Stefan Barth; Thomas Nachreiner; Christian Cremer - Web page: https://patentgrep.com/patent/US10246500B2 ## CPC - C07K 14/515: Angiogenesic factors; Angiogenin - A61K 38/1891: {Angiogenesic factors; Angiogenin} - C07K 16/28: against receptors, cell surface antigens or cell surface determinants - C07K 16/283: {against Fc-receptors, e.g. CD16, CD32, CD64 (CD23 C07K16/2851)} - A61K 38/00: Medicinal preparations containing peptides (peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00) - C07K 2317/622: Single chain antibody (scFv) - C07K 2319/00: Fusion polypeptide - C07K 2319/74: containing a fusion for binding to a cell surface receptor - C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00) - A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D) ## Abstract The technology provided herein relates to novel human cytolytic fusion proteins (hCFPs) suitable to induce apoptosis in human cells comprising a target cell-specific binding component and a human effector domain, wherein the binding component comprises an antibody or an antibody fragment with an antigen-binding site for binding to the cellular surface receptor CD64 and the effector domain comprises a variant of wild type human angiogenin (Ang) or a functional fragment thereof; to nucleic acid molecules encoding said recombinant hCFPs, vectors and host cells containing said nucleic acids and methods for preparation and producing these hCFPs. ## Claims ### Claim 1 A recombinant human cytolytic fusion protein (hCFP) suitable to induce apoptosis in human target cells, comprising a target cell-specific binding component and a human effector domain, wherein the binding component comprises an antibody or an antibody fragment with an antigen-binding site for binding to the cellular surface receptor CD64, and the effector domain comprises a variant or a functional fragment of wild type human angiogenin (Ang) as shown in SEQ ID NO: 1, wherein the variant of the wild type human angiogenin as shown in SEQ ID NO: 1 contains at least substitutions at positions that correspond structurally or by amino acid sequence homology to positions 85, 86 and 117, and wherein the substitutions are G85R, G86R and Q117G. ### Claim 2 (depends on 1) The recombinant human cytolytic fusion protein (hCFP) according to claim 1, wherein the antibody fragment is selected from the group consisting of a Fab, a scFv, a single domain, a bis scFv, a Fab2, a Fab3, a minibody, a diabody, a triplebody, a tetrabody, and a tandab. ### Claim 3 (depends on 1) The recombinant human cytolytic fusion protein (hCFP) according to claim 1, wherein the binding component is a CD64-specific single-chain variable fragment (scFv). ### Claim 4 (depends on 1) The recombinant human cytolytic fusion protein (hCFP) according to claim 1, wherein the binding component is the CD64-specific single-chain variable fragment H22(scFv). ### Claim 5 (depends on 1) A drug comprising the recombinant human cytolytic fusion protein (hCFP) according to claim 1 in combination with a pharmacologically acceptable carrier. ### Claim 6 (depends on 1) An isolated nucleic acid molecule encoding the recombinant human cytolytic fusion protein (hCFP) according to claim 1. ### Claim 7 (depends on 6) A vector comprising the nucleic acid molecule claim 6. ### Claim 8 (depends on 7) A host cell transformed with the vector of claim 7. ### Claim 9 (depends on 8) The host cell according to claim 8, wherein the host cell is a HEK293T cell. ### Claim 10 (depends on 8) A method for preparing a recombinant human cytolytic fusion protein (hCFP), the method comprising culturing the host cell of claim 8 and isolating the fusion protein from the cell culture. ### Claim 11 A recombinant human cytolytic fusion protein (hCFP) suitable to induce apoptosis in human target cells, comprising a target cell-specific binding component and a human effector domain, wherein the binding component comprises an antibody or an antibody fragment with an antigen-binding site for binding to the cellular surface receptor CD64, and the effector domain comprises a variant or a functional fragment of wild type human angiogenin (Ang) as shown in SEQ ID NO: 1, wherein the amino acid sequence of the variant comprises at least one variation as compared with SEQ ID NO: 1, wherein the variant of the wild type human angiogenin comprises the substitutions G85R, G86R and Q117G, and wherein the amino acid sequence of the variant has a sequence identify of at least 80% to SEQ ID NO: 1. ### Claim 12 (depends on 11) An isolated nucleic acid molecule encoding the recombinant human cytolytic fusion protein (hCFP) according to claim 11. ### Claim 13 (depends on 11) A vector comprising a nucleic acid molecule encoding the recombinant human cytolytic fusion protein (hCFP) according to claim 11. ### Claim 14 A recombinant human cytolytic fusion protein (hCFP) suitable to induce apoptosis in human target cells, comprising a target cell-specific binding component and a human effector domain, wherein the binding component comprises an antibody or an antibody fragment with an antigen-binding site for binding to the cellular surface receptor CD64, and the effector domain comprises a variant or a functional fragment of wild type human angiogenin (Ang) as shown in SEQ ID NO: 1, wherein the human cytolytic fusion protein (hCFP) comprises an amino acid sequence of SEQ ID NO: 3 or SEQ ID NO: 5. ### Claim 15 (depends on 14) An isolated nucleic acid molecule encoding the recombinant human cytolytic fusion protein (hCFP) according to claim 14. ### Claim 16 (depends on 14) A vector comprising a nucleic acid molecule encoding the recombinant human cytolytic fusion protein (hCFP) according to claim 14. ### Claim 17 (depends on 1) A method of treating a malignant disease, the method comprising administering an effective amount of the recombinant human cytolytic fusion protein (hCFP) according to claim 1 to a patient in need thereof. ### Claim 18 (depends on 1) A method of treating an autoimmune disease, the method comprising administering an effective amount of the recombinant human cytolytic fusion protein (hCFP) according to claim 1 to a patient in need thereof. ### Claim 19 (depends on 1) A method of a treating tissue rejection reaction, the method comprising administering an effective amount of the recombinant human cytolytic fusion protein (hCFP) according to claim 1 to a patient in need thereof. ### Claim 20 (depends on 1) A method of treating a chronic inflammatory disease, the method comprising administering an effective amount of the recombinant human cytolytic fusion protein (hCFP) according to claim 1 to a patient in need thereof.